Scientists at the John Innes Centre (JIC) Norwich, have been working for many years on nitrogen fixation, the natural process that microbes use to trap nitrogen gas and convert it into a form that they, and plants, can use. The JIC team have today, announced a discovery which adds to our understanding of how microbes sense environmental conditions are right for them to begin fixing nitrogen.

“Bacteria that fix nitrogen only do so when they sense that there is very little nitrogen available in their environment,” says Professor Ray Dixon (Project Leader at the JIC. “Normally the genes for nitrogen fixation are locked off and only unlocked and used when nitrogen levels in the environment fall. We have discovered a key piece of biochemistry that allows us to better understand how the lock operates and so may allow us to alter how it works”.

The bacterium Azotobacter vinelandii is able to fix atmospheric nitrogen when available nitrogen in its environment falls below a threshold level. Nitrogen fixation requires a great deal of energy and so the genes that carry out nitrogen fixation (so called nif genes) are tightly regulated and switched off when not required.

The nif genes are regulated by the action of two proteins, called NifL and NifA. NifA stimulates the activity of nif genes, while NifL normally binds to NifA and renders it inactive. Thus whether the nif genes are active or not depends on the interaction between these two proteins. Both proteins are sensitive to biochemical signals that occur in the bacterial cell when conditions are right for nitrogen fixation. The proteins’ physical shape and structure alters in response to these signals and this affects their ability to bind to one another. The result is that, when conditions are right for nitrogen fixation, NifA is released from the grip of NifL and is then able to stimulate the activity of the nif genes and so switches on nitrogen fixation by the cell.

The latest research has identified a single amino acid change in the NifL protein that prevents the molecule releasing NifA when the appropriate signals are present [1]. This gives the scientists an important clue about the key processes which operate the lock that controls nitrogen fixation.

The discovery will be reported in the international science journal Proceedings of the National Academy of Sciences US, and is available on line in the PNAS Online Early Edition [2].

Notes:

[1] NifA is a sigma factor dependent transcriptional activator that stimulate nif gene activity. Its action is blocked by protein-protein binding with NifL, an anti-activator. NifL is sensitive to  the  redox and fixed nitrogen status of the cell. Binding of 2-oxoglutarate (an indicator of cell carbon status) to NifA prevents NifL from inhibiting NifA . A critical arginine residue (R306) has been identified in NifL that is required to release NifA under appropriate environmental conditions. Mutation of this residue blocks release of NifA from NifL. The substitution of this arginine significantly alters the conformation of the NifL molecule and inhibits NifA’s response to 2-oxoglutarate. It appears that arginine 306 is critical for coupling the response of NifL to the cellular redox and fixed nitrogen status to a conformational switch that prevents NifL from inhibiting NifA under conditions suitable for nitrogen fixation.

[2] A crucial arginine residue is required for a conformational switch in NifL to regulate nitrogen fixation in Azotobacter vinelandii. I. Martinez-Argudo, R. Little and R. Dixon.   Article #04-05312

The John Innes Centre (JIC), Norwich, UK is an independent, world-leading research centre in plant and microbial sciences.  The JIC has over 850 staff and students.  JIC carries out high quality fundamental, strategic and applied research to understand how plants and microbes work at the molecular, cellular and genetic levels.  The JIC also trains scientists and students, collaborates with many other research laboratories and communicates its science to end-users and the general public.  The JIC is grant-aided by the Biotechnology and Biological Sciences Research Council.